Aging is a progressive accumulation of metabolic and physiologic changes associated with an increasing susceptibility to disease. Several explanations for the aging process are described in the contemporary literature. Among the most prominent is the dysdifferentiation hypothesis of aging and the membrane hypothesis of aging. The dysdifferentiation hypothesis proposes that aging is the result of a continued programmed differentiation leading to either a cessation of normal gene activity or a systematic activation of genes whose effects are deleterious to cellular function. Conversely, the membrane hypothesis of aging (MHA) states that aging is related to decreasing effectiveness of cellular protective and reparative mechanisms secondary to damage from oxygen radicals. This yields biochemical and metabolic errors, which progressively accumulate, resulting in cell aging and ultimately death. Therefore the MHA suggests that reactive oxygen metabolite (ROM) induced cell membrane structural damage is the primary mediator in cellular aging.
Reactive oxygen metabolites, also known as free oxygen radicals (FOR) are the putative initiators in the membrane hypothesis of aging. ROMs are a normal by-product of oxidative phosphorylation, and are also formed under conditions of ischemia, hypoperfusion and because of environmental contaminants. Among the many detrimental activities of ROM, or free oxygen radicals, is direct damage to mitochondrial DNA (mtDNA). Progressive accumulation of mtDNA damage renders cells unable to conduct oxidative phosphorylation reactions effectively, thereby leading to a bioenergetically deficient cell. Over time, mitochondrial DNA damage accumulates and leads to cellular dysfunction with subsequent organ failure, aging and ultimately death. This sequence forms the basis of the MHA. Additionally, there is evidence of a reduction in the oxidant-protective enzymes superoxide dis-mutase and catalase associated with aging. Thus not only are there increases in the deleterious effects of ROMs, but there is a reduction in the enzymes and mitochondrial metabolites necessary for protection from ROM and for effective mitochondrial function.
There is increasing support in the literature that reactive oxygen metabolites and mitochondrial dysfunction may initiate processes that lead to Alzheimer's disease, non-specific dementias and cognitive disorders. Thus, compounds that increase mitochondrial function and scavenge or block the activities of reactive oxygen metabolites may slow or even reverse the processes of these disorders.
Variation in life spans of certain mammalian and plant species is in part dependent upon free radical scavenging systems. It has been observed that animals with longer life spans had higher levels of SOD activity. Many studies have investigated dietary increases in antioxidants and have demonstrated life span increases by 13-30%.
.alpha.-lipoic acid
.alpha.-lipoic acid is a coenzyme for the pyruvate dehydrogenase complex in the mitochondrial matrix. It is an essential cofactor for metabolism in .alpha.-ketoacid dehydrogenase reactions. This vitamin-like substance has been supplemented orally for health benefits and has also been used as a therapeutic agent in a variety of hepatic and neurological disorders, as well as mushroom poisoning. Consideration has also been given to the use of .alpha.-lipoic acid in the treatment of diabetes mellitus and atherosclerosis, in which decreased levels of .alpha.-lipoic acid have been found. Interestingly, a specific 10.4 kb mitochondrial DNA deletion has been found in patients with diabetes mellitus and sensorineural hearing loss. Thus, it may also be hypothesized that patients with these disorders as well as aging might benefit from a diet supplemented with lipoic acid. Dietary supplementation of .alpha.-lipoic acid successfully prevents myocardial damage induced by ischemia-reperfusion injury. Presently its primary therapeutic use is for the treatment of diabetic polyneuropathy.
In physiological systems, .alpha.-lipoic acid usually exists as lipoamide covalently attached to lysine residue of the enzyme complexes. It functions in the transfer of the two-carbon fragment from .alpha.-hydroxyethylthiamin pyrophosphate to acetyl-CoA, and it gets reduced in the process. The reduced form of .alpha.-lipoic acid is dihydrolipoic acid (DHLA) containing a disulfihydral structure. DHLA has been found to exert some antioxidant actions. It has been shown to prevent microsomal lipid peroxidation by reducing glutathione which in turn recycles vitamin E. DHLA has also been demonstrated to be a free oxygen radical scavenger to reduce peroxyl, ascorbyl and chromanoxyl radicals, and to inhibit singlet oxygen.
Acetyl L-carnitine
Acetyl L-carnitine is the acetyl ester of carnitine, a biological compound which plays a key role in the transport of fatty acids from the cytosol into the mitochondrial matrix of B-oxidation. Acetyl L-carnitine modulates, through regulation of acetyl Co-A intracellular concentration, the metabolisms of sugars, lipids and amino acids, this way playing a pivotal role in cellular energy and turn over of cell membranes and proteins. Observations demonstrating a positive effect on survival were recently made on some rat populations treated with acetyl L-carnitine. The mechanisms of such an effect are currently unknown, but they are very similar to the survival observed in rats fed a restricted caloric diet. Chronic treatment with acetyl L-carnitine has shown enhanced stimulation of antiperoxidative systems, antagonism of the age-related effect on glucocorticoid secretion, increase in acetylcholine release and improvement in learning and memory. It has been reported that aged rat brain and heart possessed a reduced steady state level of mitochondrial transcripts due to reduced RNA synthesis. Pretreatment of senescent rats with acetyl L-carnitine was able to bring back the levels of mitochondrial transcripts to adult levels in a time and dose-dependent function.
Acetyl L-carnitine is capable of restoring the integrity of the cardiac mitochondrial membrane altered by aging (specifically the cardiolipin content), thereby restoring the normal activity of cytochrome oxidase, adenine nucleotide translocase, and phosphate carrier. This allows more efficient oxidative phosphorylation, and therefore, improves cardiac performance in aged animals.
Coenzyme Q-10
Coenzyme Q-10 (CoQ-10) is an important mitochondrial precursor and is essential in the transduction of energy. Reduced CoQ-10 functions as an antioxidant and can therefore combat the production of free oxygen radicals. There is evidence supporting an age-related decline of CoQ-10 in humans and other species, thus further supporting the membrane hypothesis of aging. CoQ-10 is currently used alone or in combination as a health/nutritional supplement. However, there is evidence suggesting that long-term supplementation of this precursor alone has no beneficial effect on lifespan or energy production. However, it is believed that when combined with acetyl L-carnitine and lipoic acid, cellular degeneration is limited and energy production is enhanced.
Glutathione
Glutathione (L-gamma-glutamyl-L-cysteinyl-glycine; GSH) is an endogenous thiol that detoxifies reactive oxygen species. It is also involved in the metabolism and detoxification of xenobiotics, drugs and their metabolites and offers protection from oxidizing ROM via reactions catalyzed by GSH-S-transferase, transpeptidases, transhydrogenases, and peroxidases and reductases. Mitochondrial glutathione is critical to cell viability, and the glutathione redox cycle is a primary antioxidant defense system within the mitochondrial matrix. Additional functions include intracellular binding, transport of lipophilic substances and prostaglandin synthesis.
Many studies have demonstrated that alterations of glutathione levels through excess or reduced production have a beneficial or harmful influence in cellular function, respectively. Specifically, it has been shown that sulfhydryl compounds reduce gentamicin induced outer hair cell damage in vitro and that gentamicin ototxicity, in vivo, may be reduced using glutathione, and diethyldithiocarbamate offers protection from cisplatin ototoxicity. Conversely, systemic inhibition of glutathione synthesis potentiates the ototoxicity of a combination of ethacrynic acid and kanamycin, and cis-platin rephrotoxicity is potentiated by GSH depletion. It has also been shown that the mechanism for toxicity of certain clinically used drugs occurs secondary to reduced glutathione levels with an increase in reactive oxygen species. Recent studies have demonstrated an 86% age-associated reduction in glutathione levels in the auditory nerve while other cochlear tissues have had stable levels.
Studies of patients with Alzheimer's disease have demonstrated age-dependent decreases of glutathione-peroxidase activities and their cofactors. Additionally, there was a significant increase in plasma glutathione peroxidase which was interpreted as an imbalance between reactive oxygen species and peripheral antioxidant opposing forces. These findings were not observed in controls. Other enzyme changes have been studied, including glutathione reductase, catalase and superoxide dismutase in patients with Alzheimer's disease. The findings demonstrated significantly elevated enzyme activity levels where lipid peroxidation was most prevelent, as documented with levels of thiobarbituric acid. It is believed that there is a compensatory elevation in antioxidant activity in response to increased free radical formation.
Piracetam
Piracetam (C.sub.6 H.sub.10 N.sub.2 O.sub.2) is a nootropic compound which exhibits cognition-enhancing properties possibly through a potentiation of neurotransmission, although the formal mechanism has not been established. Piracetam possesses a very low toxicity and lacks serious side-effects. This compound has been used in several clinical and experimental conditions including: the treatment of motion sickness, protection of hypoxia-induced amnesia and enhancement of acquisition in passive avoidance experiments with rats. In general, piracetam seems to be somewhat effective in patients with mild to moderate dementia, Alzheimer's disease, Parkinson's disease and in the psychotic state schizophrenics. Additionally, complete recovery of sudden deafness has been shown with piracetam. Piracetam demonstrates an ability to increase systemic and microvascular circulation possibly because of decreased platelet aggregation, enhancement of red blood cell deformability and adherence of erythrocytes to endothelial cells. Piracetam has been shown to increase the survival rate of rats subjected to hypoxia and also to decrease the time of recovery from hypoxia. Piracetam is absorbed very well after oral administration with a 100% bioavailability. It is excreted practically unchanged in the urine and completely eliminated after thirty hours.
These compounds together will synergistically act against the processes of cellular degradation through their antioxidant properties and their ability to upregulate mitochondrial function, as well as other physiologic and biochemical actions. The nutritional supplement of the present invention overcomes the limitations of the prior art in that it utilizes the synergistic combination of nutritionally effective amounts of the above-referenced compounds to enhance mitochondrial function. These and other advantages of the present invention will be readily apparent from the description, discussion and experimental examples which follow.